This invention is termed a preliophic moleculator and process for moleculation as an abbreviated terminology for a protonic-electronic-ionic-photonic molecular computer. Preliophic is derived from the first two letters of words in the hyphenated description along with the last “ic”, thereby creating the coined term. The preliophic molecular computer is a device used to manipulate the motion and interaction of selected molecules in a medium (e.g., polyacrylamide, agarose or starch gel) under the influence of an electric field, induced pH gradient, or by some other means. The device and process are invented to mimic features of inferred interactions of molecules in cellular structures. However, it is expected that the physical device has application beyond experimental studies in molecular and cell biology, and in producing molecular byproducts. It will comprise a useful tool in all fields of chemistry, including theoretical, conceptual, logical and mathematical studies involving molecular interactions in a controlled environment. Indeed, the conceptual embodiment of the physical device and process are central elements in several derivative novel, logical, albeit counterintuitive (i.e., non-obvious) claims. These include: vaccines for use in acquired immunodeficiency syndrome (AIDS) and other lentivirus-related infections involving single or multivalent killed vaccines against relatively uncommon pathogens, though not against the lentiviruses themselves; devices used in elucidating “autotoxicity,” “autovirulence,” and “context specificity”; conceptualizing a gedanken study designed to reveal both changes in DNA in brain and measure changing G*C::A*T ratios using single-nucleotide polymorphism (SNP) analyses involving DNA extracted from cells in a hair follicle and three sections in brain; and, non-invasive methods, approaches and technologies for measuring nurture quantitatively, in both living and artificial environments, by using G*C::A*T ratios. Each of these corollary inventions is claimed in this patent application.
Molecular interactions produced in preliophic moleculation processes and devices are a direct contrast to the lack of molecular interactions in electrophoresis and electrophoresis devices. That is, electrophoresis (in contrast to preliophic moleculation) facilitates parallel movements of molecules which, by definition of parallelism, cannot interact. Thus, the preliophic moleculator and its processes are useful in elucidating and disambiguating molecular function, whereas electrophoresis is useful in elucidating molecular structure.
Typically, the medium used with the preliophic moleculator device and process is a gel or substrate similar to types used in electrophoresis devices and processes, and especially medium used in isoelectric focusing (IEF). However, it is expected that other non-gel mediums also may prove useful (e.g., viscous fluids, polyacrylamides, or other compositions through which intact, non-denatured molecules are able to migrate under the influence of electric fields or gradients). In this description such substances are termed substrate mediums. (NB: Throughout this description, the term “mediums” deliberately is used as the plural of medium, to distinguish mediums from media.)
The impetus for this invention is a series of discoveries and inferences suggesting that parsimony exists among most, if not all, molecular mechanisms of long-term memories (LTM) in living systems accounting for evolution, speciation, development, differentiation, immunity, cognition and behavior, aging, and death. Central to that parsimony are changes in DNA, claims to the contrary notwithstanding. As examples, it generally is agreed that “selection” among random changes in DNA is a basis of Darwinian evolution. Barbara McClintock generally is credited with demonstrating that changes in DNA (dubbed transpositions or “transposons”) account for the developmental process producing variegation in maize. Other changes in and to DNA (including methylation) are associated with development and differentiation from singular cell to multiple cell organisms. Susumu Tonegawa demonstrated that rearrangements in DNA are central to immune memory and recognition. Compelling evidence now suggests that LTM in brain must involve a priori changes in DNA, especially in non-proteomic regions of the genome, with subsequent a posteriori generation and development of axons and dendrites in the neural network.
Changes in DNA in brain differ fundamentally from changes in DNA described by McClintock and Tonegawa insofar as changes in DNA in brain are not accompanied by cell division. Evolutionarily, it is thought that the bony cranium served to constrain or ablate cell division, while retaining molecular mechanisms for changing DNA especially in brain. Equally important, cell division in brain would be counterproductive because of the information redundancy. In other words, redundancy derived from cell division and clonal expansion in the immune system is desirable, whereas the lack of cellular redundancy (albeit with increased neuronal axon and dendrite connectivity) in brain, serves to facilitate molecular information processing in neurons. In the end, those DNA changes in brain are not heritable and/or transmitted to germline cells (e.g., sperm and ova) thereby differing from both Darwinian and Lamarckian evolution. Indeed, a major reason for inventing the preliophic device and process is to provide a concrete embodiment of an inverse to Francis Crick's “central dogma.” According to the “central dogma” molecular information generally flows from DNA to RNA (by a process called transcription), and from RNA to proteins (by a process called translation). Crick subsequently allowed for “reverse transcription” after the discovery of retroviruses, yet it is noteworthy that the “central dogma” was flawed at the outset. For this thesis and invention to be viable, there are no “dogma” regarding movements of molecules into, within and out of living cells beyond being directed and vectorial, and in a manner that permits interactions among other directed and vectorial molecules.
Four key, albeit unreported and non-obvious, observations provided an additional logical basis for this invention. First, although there is a well established chemio-osmotic hypothesis for energy production and electron transfer within mitochondria in cells, a purpose for hydrogen ion (i.e., proton) byproducts pumped from mitochondria was never fully elaborated. Second, microheterogeneous commercial amphoteric molecules (i.e., “ampholytes”) used in IEF and other electrophoretic processes are remarkably analogous to and concordant with the microheterogeneity in natural intercellular microtubulin-associated proteins (also known as “microtubule associated proteins,” [MAPs]), and phosphatidyl-proteins and other phosphatidyl-moieties associated with cell membranes (e.g., phosphatidylserine, phosphatidylinositol, phosphatidylinesitol, phosphatidylglycerol, phosphatidylethanolamine, phosphatidylethylamine). Third, the geometry of all electrophoretic devices, whether in one- or two-dimensional electrophoresis activities, is designed to examine and determine molecular structure based on parallel-movements of molecules in electric fields or proton gradients, with there being no potential for those devices to examine and determine molecular function. Structure is elucidated because molecular species move in parallel, with molecular weight and charge often serving to define structure. Fourth, cellular histology, a vast variety of reported and unreported clinical, pathological and experimental findings, and preliminary experimentation, all point to the logic of a “molecular computer” based on molecules moving toward their isoelectric points (pl), though interacting with other molecules moving toward their isoelectric points.
The invention reported here is initially comprised of a circular device supporting a type of IEF medium, typically a gel with one charged pole in the center and an oppositely charged pole, comprises the entire perimeter of the device. For example, with a positive pole at the center and the perimeter serving as the negative pole, electronegatively charged molecules will move toward the center and electropositively charged molecules will move towards the cell boundary (i.e., perimeter). This is precisely as it happens in cells wherein basic molecules generally move toward the perimeter of the cell where the pH is approximately 7.4, and acidic molecules generally move toward the center of the cell where the pH can be as low as 2 to 3. Even without its actual construction, this conceptual embodiment of a preliophic moleculator now provides a conceptual basis for cellular micro-geography thereby providing logical and theoretical reasons for the nucleus generally being situated in the center of cells where pH values are low, and other cellular structures being distributed more peripherally in cells, regardless of cell shapes. (NB: The term pH is a designation for the percentage of hydrogen ions. Also noteworthy is that shapes other than circular shapes work equally well in this invention, including triangles, pie-shapes, etc.).
A temperature regulated system is incorporated in the device both to accommodate the dissipation of heat and to regulate the rate molecules may move in their directed pathways. Not only does the temperature regulated system obviate denaturing molecules, it facilitates modeling cold- versus warm-blooded molecular information processing.
The terms, “moleculator” and “moleculation” are used, respectively, to define the device and process for inducing the controlled migration of molecules in a substrate medium under the influence of an electric field, pH-gradient or other means. Implied in the terminology is the study of the interaction of migrating molecules, or molecules that approach or have reached their isoelectric location. Also implied in the terminology is the study of arbitrarily close interacting molecules, whether these interactions take place at some pH or otherwise.
The design of the device is particularly adapted for the study of biological functions and processes, as the circular configuration emulates the generally spherical configuration of the biological cell. Avenues for studying the dynamics involved in the progression or expression of disease also are provided by use of the moleculator device. For example, although rarely discussed and despite its ubiquity, the Epstein-Barr virus (EBV) is associated with more than 92 different diseases and syndromes, some of which involve “hit-and-run” and/or “beneath-the-radar” pathology. Many of these 92 diseases and syndromes mimic or comprise psychosomatic illnesses that often are stress-induced. The intriguing question was why would a common and ubiquitous virus be associated with such a wide variety of diseases and syndromes, yet almost never do more than two of these 92+ diseases or syndromes occur in the same person? What possible molecular mechanisms could account for this counterintuitive finding and seeming conundrum? What theories and technologies could answer these questions?
The terms “autotoxicity,” “autovirulence” and “context-specificity” were coined in 1983 to explain these and other phenomena, including prion-related diseases. A prototype of autovirulence is specifically implicated in stress-induced EBV-associated consequences in HIV and AIDS (e.g., dose-dependent production of acid-labile alpha-interferon). There was other evidence of EBV-induced aberrant translation products associated with the EBER I and EBER II small nuclear ribonucleoproteins (snRNPS) [RNA plus protein particles]. Other viruses and their snRNPs also could contribute to the production of aberrant translation products (e.g., snRNPs comprising VA I and VA II associated with some adenoviruses), whereas lentiviruses (e.g., HIV) are more likely to be associated with aberrant transcription products. Finally, it was noted that extant theories of autoimmune diseases (e.g., that the immune system is abnormally reacting to “self”) could give way to novel possibilities that “self” or “abnormal-self” are abnormally presented to a healthy immune system, possibly involving “molecular mimicry” of “self.” What remained was to invent an appropriate and practical technology to explicate these theories, findings and other phenomena.
Evidence of EBV-associated aberrant translation products in a variety of EBV-associated diseases and syndromes was reported. Inferential evidence suggests EBER I and/or EBER II contribute to aberrant translation products and molecular mimicry. The preliophic molecular of this invention is designed to study molecular function and infer molecular mimicry experimentally, even in the absence of precise details of amino-acid sequences, conformational variations and/or molecular mechanisms. It is suggested that the etiology of EBV-associated diseases and syndromes ultimately can be unraveled with the aid of the preliophic molecular.
It is noteworthy that preliophic moleculation is unlike electrophoresis in at least one very significant regard. As noted, electrophoresis provides a simple, excellent and economical approach for purifying structure. By way of contrast, arbitrarily complex functional systems can be elucidated and explicated using preliophic moleculators. For example, the molecular biology, genetics and genomics of long-term memory (LTM) in living systems can be studied using the preliophic moleculator of this invention, and in some instance providing logical or mathematical proofs, against claims to the contrary notwithstanding.
A 1979 review by one of the inventors of six decades of research on molecular bases of LTM in living systems supports changes in DNA as a priori bases for LTM in brain, with axon-dendrite connectivity representing a posteriori bases for neural networks. Furthermore, no models of LTM are mathematically or logically complete without affirming or rejecting the claim that “the definitive evaluation of DNA as the ultimate repository of information” in brain (Smith 1979). The 1979 review then cited six novel lines of inquiry supporting the DNA change hypothesis. Two decades later, research involving five of six lines of inquiry now provides further evidence supporting DNA changes as a basis of LTM. Insofar as the device is designed to emulate molecular information flow at the cellular level, the preliophic moleculator now provides a means to examine a corollary to the remaining line of inquiry. In the end, preliophic moleculation enables one to model changing G*C::A*T ratios over time, both experimentally and concretely. At a practical level, this line of inquiry also implicates host mechanisms contributing to the microheterogeneity in lentiviruses, thereby contributing to the “proof” that neutralizing vaccines cannot be produced against HIV or other lentiviruses; that is, the intrinsic unreliability of reverse transcriptases and host factors contribute to lentivirus microheterogeneity. Inferentially, the conceptual embodiment of the preliophic moleculator reveals possibilities for non-invasive imaging technologies to model changing G*C::A*T ratios over time in intact cells and organisms, and in advanced preliophic devices. Such non-invasive imaging technologies most likely will focus on spin properties of phosphorus (in contrast to hydrogen) atoms in nuclear magnetic resonance studies.
If DNA in brain is subject to changes in association with LTM, it stands to reason that Francis Crick's “Central Dogma” of molecular biology is inadequate for explaining a necessary inverse flow of information possibly based on the input of conformational or other non-sequential representations of molecular information in contrast to sequential information embodied in the “central dogma.” Interestingly, Crick may have anticipated this shortcoming given his little noted, albeit extraordinary, comment in a footnote: “There is, for example, the problem of the chemical nature of the agent of the disease scrapie.” Nevertheless, we know enough to say that a non-trivial example showing that the classification was wrong could be an important discovery.” This comment suggests a logical corollary about the flow of information in cells. Regardless, both pathways of information transfer [DNA to RNA to Proteins (connoted the “Central Dogma”)] and an inverse pathway (conformational molecular information to endogenous- or exogenous-RNA-mediated reverse transcription (or other possible novel mechanisms) to changes in largely non-protein encoding regions of DNA) now can be studied in the invented preliophic moleculating device that emulates cellular molecular information processing (i.e., “moleculations”). Although there is not a Latinate term of art for this emulation process, preliophic molecular processes fall somewhere between being in vitro and in vivo. Indeed, preliophic moleculation, through its capture of functional molecular information processing, now makes it possible to educe mathematical and logical proofs that would not be possible in vivo and in vitro. The term “biological and chemical computability” is used to capture these logical and mathematical proofs, much akin and analogous to the notion of computability in computer sciences. The difference is that biological and chemical computability requires additional information about space, time, temperature and molecular vectorial positioning. As previously noted, this device and its moleculation processes also emulates artificial and cellular micro-geography positioned between in vivo and in vitro studies.
In the study of LTM and the evaluation of DNA as the ultimate repository of “information” (i.e., LTM) in the brain, the preliophic moleculator can aid in the empirical measurement and determination of DNA changes in the brain. The study of post-mortem brain tissue of exceptionally gifted individuals, descendants of such individuals at different ages, sets of identical twins raised together and apart, different sections of individual brains, and other such comparative studies, can be accomplished using the device of this invention. The potential for nuclear magnetic resonance studies already was cited. Use of fluorescent or other dyes also could find application, particularly in assessing changing G*C::A*T ratios.
Another complex conceptual embodiment of preliophic moleculation involves the study of causality in AIDS, and the formulation of vaccines for use in treating AIDS and other lentivirus infections. The former is mostly theoretical and inferential; the latter is largely empirical, though based on complex elements of the established cause. A brief sketch of the underlying reasoning follows.
Lentiviruses are one of three classes of retroviruses that produce changes in DNA when retrovirus RNA is “reverse transcribed” to form a DNA “provirus.” A unique feature of lentiviruses—supported by a variety of clinical, experimental and epidemiological data—is that their proviruses generally occupy protected subclasses of cells in the immune system having LTM potential, which then are trans-activated by relatively uncommon nascent pathogens, though not relatively common pathogens. Other classes of retroviruses (i.e., oncoviruses and spumaviruses) generally lack trans-activation mechanisms, and generally do not occupy protected classes of immune system cells having LTM potential.
Although lentivirus trans-activation can be recursive in intact systems (i.e., lentiviruses can trans-activate themselves), lentivirus trans-activation most often is caused by relatively uncommon pathogens—so called opportunistic pathogens. Although distinctions between common and uncommon pathogens in AIDS have been known since the early 1980s, reasons for these distinctions remained elusive until the advent of conceptual preliophic moleculators.
Lentiviruses now provide a convenient means for highlighting several advanced preliophic moleculation applications possibly requiring multiple (and/or networked) preliophic devices. One application is to establish the cause of AIDS. To many scholars and investigators, HIV is the sine qua non for AIDS, whereas to others HIV may play no role in AIDS. Whether HIV is a necessary and sufficient element in AIDS, and the general matter of causality in AIDS has never been addressed; again, claims by O'Brien and Goedert (1996) to the contrary notwithstanding. Perhaps more broadly, are there necessary and sufficient rules or conditions required to establish causality in most infectious diseases? What roles can preliophic processes play in disambiguating these issues?
Among early attempts at defining conditions to establish causality are studies of anthrax by Robert Koch, based on work with his mentor, Jacob Henle. Koch formulated a set of postulates, often referred to as the Henle-Koch Postulates (HKP), which he used in demonstrating that Bacillus anthracis is indeed the cause of anthrax. Four elements are central to Koch's “proof.” First the pathogen must be isolated from the intact organism. Second, the isolated pathogen must be inoculated into a single cell-type, which, in turn, produces characteristic pathological changes. Third, the isolated pathogen must be transmitted to an animal model, causing like disease. Fourth, once isolated again from the animal model, the pathogen must be inoculated in species from which it was first inoculated, and then shown to cause the same underlying disease. A critical theme in these postulates is that a single cell-type links the pathogen to the disease. An underlying axiom from sentential logic captures the spirit of these HKP; to with, Modus Penendo Ponens (if “A implies B” is true; and, if “A” is true; then, “B” is true). That is, if Bacillus anthracis causes anthrax is true; and, if Bacillus anthracis is the true pathogen associated with anthrax; then, anthrax can be inferred (to be the true consequence of Bacillus anthracis).
If HIV and other lentivirus infections are to be shown to be causes of AIDS or other diseases using HKP, a single cell-type must be associated with disease. Not only is this not true based on clinical, laboratory, experimental and epidemiological evidence, preliophic moleculators bring this into sharper focus. Multiple cell-types are associated with diseases caused by relatively uncommon (nascent) “opportunistic” pathogens that trans-activate the HIV (or other lentivirus) provirus in a separate cell-type. Advanced preliophic moleculation processes will emulate networks of multiple preliophic moleculators—one set perhaps to emulate the consequences of trans-activation, and other sets perhaps to emulate associations between opportunistic pathogens and diseases.
Because HIV cannot be shown to be the sole or sufficient cause of AIDS using the HKP, what recourse is there? Proof lies in understanding distinctions between the effects of relatively common and uncommon pathogens in the HIV-infected individual (or in other lentivirus infections). There are compelling clinical, laboratory and epidemiological data that suggest that AIDS is not associated with relatively commonplace pathogens in HIV-infected persons. In short, a category of pathogens—so called relatively commonplace pathogens—do not play a role in trans-activating HIV. Indeed, considerable evidence suggests that the immune system remains intact for those pathogens. This finding can be emulated and modeled using preliophic moleculators, and give rise to applications of a second axiom from sentential logic; to wit, Modus Tollendo Tollens (if “A implies B” is true; and, if “not B” is true; then, “not A” is true). Modus Tollendo Tollens sometimes is referred to as “the rule of denial.” Application of this axiom, when B comprises the relatively commonplace pathogens in an environment, confirms clinical, epidemiologic and laboratory findings; to wit, the cause of AIDS is HIV and selected relatively rare opportunistic pathogens. Although initially counterintuitive, an obvious corollary is that multivalent killed vaccines against relatively uncommon pathogens may circumvent trans-activation of HIV and other lentiviruses. This finding is affirmed in naturalistic studies involving Icelandic sheep between 1933 and 1954, simian AIDS-like diseases among primates in an animal colony at UC Davis during the late-1970s and mid-1980s, and in epidemiological data collected on human pathogens worldwide.